GSM6022405: SI Crypts_sc_WT3; Mus musculus; RNA-Seq
Small intestine crypts were isolated from three WTwt and three Tet2/3 dkoKO mice as previously described (Sato and Clevers, 2013). Single-cell suspensions from intestinal crypts were obtained by enzymatic digestion, incubating crypts in MEM medium supplemented with 2 mg/ml Trypsin and 2000 U/ml DNAse I for 30 minutes at 37ºC in a water bath, as previously described (Grun et al., 2015). Apoptotic and dead cells were removed from suspension using the Dead Cell Removal Kit (Miltenyi Biotec). Approximately 20,000 cells per sample were loaded into the Chromium Controller system, and libraries were subsequently prepared using the Chromium Single Cell 3' Reagent Kit v.2 (10X Genomics), following the manufacturer's instructions. Libraries were sequenced on a HiSeq 4000 device (Illumina) using paired-end (26+74bp) settings for 100 cycles.